The biologic basis to the evolution of platinum resistance has been attributed to changes in many cellular functions including drug efflux, glutathione levels, and DNA repair capacity. However, a comprehensive understanding of the global molecular changes that accompany the development of platinum-resistance in ovarian cancer cells has not yet been elucidated. Current technologies cannot efficiently determine the potential therapeutic response of a cancer prior to treatment. Platinum compounds are chemotherapeutic agents effective in treatment of many human solid tumors. Response to platinum-based chemotherapy is one of the most critical determinants of outcome for patients with advanced stage epithelial ovarian cancer. Currently the standard treatment protocol used in the initial management of such patients is primary cytoreductive surgery, followed by adjuvant therapy with a platinum and taxane. Approximately 70% of patients will have a complete clinical response to this initial therapy, with absence of clinically detectable residual disease on clinical examination, radiologic imaging, or serum CA125 tumor marker. However, for most patients, remission is short-lived, and the majority will develop recurrent disease that ultimately becomes resistant to further platinum therapy, resulting in extremely poor survival.
The BCL-2 family of proteins govern mitochondrial outer membrane permeabilization and constitute an intracellular checkpoint of apoptosis, largely defined by conserved motifs termed BCL-homology regions. (Yin et al, Nature 369:321-323, 1994 which is incorporated by reference). The BCL-homology regions 1, 2, 3 and 4 (BH1 through BH4) domains have been shown crucial for function (Yin et al. Nature 369:321-20 323, 1994 which is incorporated by reference; Boyd et al., Oncogene 11:1921-1928; Chittenden et al., Embo J 14:5589-5596, 60 1995 which are incorporated by reference). Members of the BCL-2 family typically can competitively heterodimerize and homodimerize, determining whether a cell will respond to an apoptotic signal (Oltvai and Korsmeyer, Cell 79:189-192, 1994 which is incorporated by reference).
BAD (BCL-2 Associated Death Promotor) is a proapoptotic Bcl-2 family protein that regulates the intrinsic apoptosis pathway. In its transient state, BAD is phosphorylated, rendering the protein inactive. Phosphorylated BAD interacts with 14-3-3 scaffold proteins in the cytoplasm, until cleavage by caspase-3 or dephosphorylation by calcineurin allows the release of BAD. 14-3-3 binding has been shown to be sequence-specific to a phosphoserine containing motif (Muslin et al. Cell 84:889-896, 1996 which is incorporated by reference), based on phosphorylation of serine residues (Serine-259 and Serine-621) in Raf-1. Once BAD is dephosphorylated (posttranslational modification), it is active; it translocates from the cytosol to the mitochondria and forms heterodimers with BCL proteins to block the antiapoptotic functions of the proteins.
Current technology does not monitor cellular phosphorylation status to determine the potential for platinum therapy resistance. Accordingly, there is an unmet need to develop screening systems to aid in the analysis and prognosis of current and possible future therapy resistance.